• Title/Summary/Keyword: High-level expression

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In situ Recovery of hCTLA4Ig from Suspension Cell Cultures of Oryza sativa (형질전환 벼 현탁세포 배양에서 hCTLA4Ig의 in situ 회수)

  • Choi, Hong-Yeol;Cheon, Su-Hwan;Kwon, Jun-Young;Yun, Boreum;Hong, Seok-Mi;Kim, Sun-Dal;Kim, Dong-Il
    • KSBB Journal
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    • v.31 no.4
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    • pp.284-290
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    • 2016
  • In this research, recombinant human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) was produced by transgenic rice cells. RAmy3D promoter was used for overcome the limitation of low expression level in transgenic plant cells, and the secretion of target protein was accomplished by signal peptide. However, the RAmy3D promoter system which can be induced only by sugar starvation causes the decrease of cell viability. As a result, cell death promotes the release of protease which degrades the target proteins. The protein stability and productivity can be significantly influenced by proteolysis activity. Therefore, development of new strategies are necessary for the in situ recovery of target proteins from cell culture media. In this study, in situ recovery was performed by various strategies. Direct addition of Protein A resin with nylon bag leads to cell death by increased shear stress and decrease in production of hCTLA4Ig by protease. Medium exchange through modified flask could recover hCTLA4Ig with high cell viability and low protease activity, on the other hand, the productivity was lower than that of control. When in situ recovery was conducted at day 7 after induction in air-lift bioreactor, 1.94-fold of hCTLA4Ig could be recovered compared to control culture without in situ recovery. Consequently, in situ recovery of hCTLA4Ig from transgenic rice cell culture could enhance productivity significantly and prevent degradation of target proteins effectively.

Enhancement of phagocytosis and cytotoxicity in macrophages by tumor-derived IL-18 stimulation

  • Xu, Henan;Toyota, Naoka;Xing, Yanjiang;Fujita, Yuuki;Huang, Zhijun;Touma, Maki;Wu, Qiong;Sugimoto, Kenkichi
    • BMB Reports
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    • v.47 no.5
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    • pp.286-291
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    • 2014
  • Inoculation of mice with the murine NFSA cell line caused the formation of large tumors with necrotic tumor cores. FACS analysis revealed accumulations of $CD11b^+$ cells in the tumors. Microarray analysis indicated that the NFSA cells expressed a high level of the pro-inflammatory factor interleukin-18 (il-18), which is known to play a critical role in macrophages. However, little is known about the physiological function of IL-18-stimulated macrophages. Here, we provide direct evidence that IL-18 enhances the phagocytosis of RAW264 cells and peritoneal macrophages, accompanied by the increased expression of tumor necrosis factor (tnf-${\alpha}$), interleukin-6 (il-6) and inducible nitric oxide synthase (Nos2). IL-18-stimulated RAW264 cells showed an enhanced cytotoxicity to endothelial F-2 cells via direct cell-to-cell interaction and the secretion of soluble mediators. Taken together, our results demonstrate that tumor-derived IL-18 plays an important role in the phagocytosis of macrophages and that IL-18-stimulated macrophages may damage tumor endothelial cells.

Identification and Characterization of Three Differentially Expressed Ovarian Genes Associated with Ovarian Maturation in Yesso Scallop, Patinopecten yessoensis

  • Kim, Young-Ju;Kang, Hye-Eun;Cho, Gyu-Tae;Suh, Young-Sang;Yoo, Myong-Suk;Kim, Hyun-Woo
    • Fisheries and Aquatic Sciences
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    • v.12 no.4
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    • pp.276-285
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    • 2009
  • Despite great commercial interest, relatively little has been described about molecular mechanism of bivalve reproduction. We investigated genes involved in ovarian maturation of the Yesso scallop, Patinopecten yessoensis. GSI index and histological analysis revealed that maturation of ovary begin in February and spawning period is from April to June which is similar to the previous study in the East Sea. As result of combination analysis of differential display RTPCR (DDRT-PCR) and histological examination, vitellogenin (Vg), ferritin (Ft) and ADT/ATP carrier protein (ACC) were identified as differently expressed genes in maturating ovary. Endpoint RT-PCR results showed that Vg is ovary-specific genes whereas Ft and ACC are expressed ubiquitously suggesting that Vg can be good molecular markers for ovarian development and sex determination in bivalves. Quantitative PCR results revealed that Vg were expressed highest during growth stage and appears to play a major role in oocyte maturation. On the contrary, expression of Ft was highest after spawning stage, which suggests that up-regulation may be involved in spawning and inactive stages in which the scallops recover from spawning. In addition, high level of the mitochondrial gene, ACC, may play a role in energy metabolism in maturating oocytes. Isolation and molecular studies of these key genes will expand our knowledge of the physiological changes from various exogenous factors including temperature, salinity, pH, even or numerous endocrine disrupting chemicals (EDCs) during reproductive cycle. In addition, further study of these genes implicates various industrial applications including the stable seed production, increased food quality, or economic aquaculture system.

Prediction of Non-Genotoxic Carcinogenicity Based on Genetic Profiles of Short Term Exposure Assays

  • Perez, Luis Orlando;Gonzalez-Jose, Rolando;Garcia, Pilar Peral
    • Toxicological Research
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    • v.32 no.4
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    • pp.289-300
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    • 2016
  • Non-genotoxic carcinogens are substances that induce tumorigenesis by non-mutagenic mechanisms and long term rodent bioassays are required to identify them. Recent studies have shown that transcription profiling can be applied to develop early identifiers for long term phenotypes. In this study, we used rat liver expression profiles from the NTP (National Toxicology Program, Research Triangle Park, USA) DrugMatrix Database to construct a gene classifier that can distinguish between non-genotoxic carcinogens and other chemicals. The model was based on short term exposure assays (3 days) and the training was limited to oxidative stressors, peroxisome proliferators and hormone modulators. Validation of the predictor was performed on independent toxicogenomic data (TG-GATEs, Toxicogenomics Project-Genomics Assisted Toxicity Evaluation System, Osaka, Japan). To build our model we performed Random Forests together with a recursive elimination algorithm (VarSelRF). Gene set enrichment analysis was employed for functional interpretation. A total of 770 microarrays comprising 96 different compounds were analyzed and a predictor of 54 genes was built. Prediction accuracy was 0.85 in the training set, 0.87 in the test set and increased with increasing concentration in the validation set: 0.6 at low dose, 0.7 at medium doses and 0.81 at high doses. Pathway analysis revealed gene prominence of cellular respiration, energy production and lipoprotein metabolism. The biggest target of toxicogenomics is accurately predict the toxicity of unknown drugs. In this analysis, we presented a classifier that can predict non-genotoxic carcinogenicity by using short term exposure assays. In this approach, dose level is critical when evaluating chemicals at early time points.

Comparative Proteomic Profiling of Pancreatic Ductal Adenocarcinoma Cell Lines

  • Kim, Yikwon;Han, Dohyun;Min, Hophil;Jin, Jonghwa;Yi, Eugene C.;Kim, Youngsoo
    • Molecules and Cells
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    • v.37 no.12
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    • pp.888-898
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    • 2014
  • Pancreatic cancer is one of the most fatal cancers and is associated with limited diagnostic and therapeutic modalities. Currently, gemcitabine is the only effective drug and represents the preferred first-line treatment for chemotherapy. However, a high level of intrinsic or acquired resistance of pancreatic cancer to gemcitabine can contribute to the failure of gemcitabine treatment. To investigate the underlying molecular mechanisms for gemcitabine resistance in pancreatic cancer, we performed label-free quantification of protein expression in intrinsic gemcitabine-resistant and -sensitive human pancreatic adenocarcinoma cell lines using our improved proteomic strategy, combined with filter-aided sample preparation, single-shot liquid chromatography-mass spectrometry, enhanced spectral counting, and a statistical method based on a power law global error model. We identified 1931 proteins and quantified 787 differentially expressed proteins in the BxPC3, PANC-1, and HPDE cell lines. Bioinformatics analysis identified 15 epithelial to mesenchymal transition (EMT) markers and 13 EMT-related proteins that were closely associated with drug resistance were differentially expressed. Interestingly, 8 of these proteins were involved in glutathione and cysteine/methionine metabolism. These results suggest that proteins related to the EMT and glutathione metabolism play important roles in the development of intrinsic gemcitabine resistance by pancreatic cancer cell lines.

Effects of Baedokhwanbalhyobang (BDHBH) on Immune Modulation in Dermatitis Model of NC/Nga Mice (배독환발효방(排毒丸醱酵方)의 면역조절작용을 통한 항아토피 효능)

  • Gim, Seon-Bin;Kim, Soo-Myung;Kim, Dong-Hee
    • Journal of Haehwa Medicine
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    • v.19 no.2
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    • pp.101-118
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    • 2011
  • Herbal medicine has a high body absorption rate when it ferments. Biological and clinical research on the fermented herb gradually increases because it has effective materials for the treatment of a disease and it is a little bitter. In this study, we investigated the effect of fermented Baedokhwanbalhyobang (BDHBH) on attenuation of the development of atopic dermatitis in NC/Nga mice by evaluating the cytokine level in serum, the mRNA expression of cytokine and histological alteration of the skin, and the skin severity. We have come to the following conclusion. BDHBH led to a significant decrease in the skin severity score (63.1%) as compared to the control group. CD4+/CD45+, CD4+, B220+/CD23+, and CD11b+/Gr-1+cells of peripheral mononuclear cells (PBMCs) in the BDHBH-treated group were decreased to 6.7%, 31.1%, 22.4%, 36.6%, respectively. CD3+and CD11b+/Gr-1+immune cells in dorsal skin of the BDHBH-treated group were decreased to 52.9% and 28.0%. The levels of IL-5 and IL-13 in serum of the BDHBH-treated group were inhibited to 18.8% and 5.1%. The mRNA expressions of IL-5 and IL-13 in dorsal skin were also decreased to 30.6% and 27.8% after the treatment of BDHBH. BDHBH inhibited the proliferation and differentiation of eosinophils. In histological examination, BDHBH decreased the thickness of epidermis and dermis, and infilatration of mast cells as compared to the control group. These results indicate that BDHBH inhibits the pathogenic development of atopic dermatitis in NC/Nga mice. These results may indicate that BDHBH attenuates the development of atopic dermatitis-like lesions by lowering immune cells and inflammatory cytokine levels, and that it is valuable in drug development for the treatment of atopic dermatitis. Further experiments on the components of BDHBH will be needed to better understand the effect of a fermented herb as compared to a herb.

A Study on the Effective application scheme of Subway optical communication system (지하철 광통신 시스템의 효율적 운용방안에 대한 고찰)

  • Kwon, Gi-Jung;Lee, Gi-Seung;Ko, Young-Yun
    • Proceedings of the KSR Conference
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    • 2007.11a
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    • pp.1541-1550
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    • 2007
  • A subway is one of many public transportation means that contributes toward our society including seoul and offering of its service and expression of various information are in need due to users high demand. And there are various class of users. For that reason an optical communication network in the general contorl center and station buildings was installed. Considering the importance of safe operation of subway, optical communication exists to enhance customer satisfaction. Safe operation is essential as its level of reliance accounts for about 95%. this paper will look for ways to promote mutual information interchange and operation know-how with systems of the same kind and sharing a capable of effective operation of systems in place. Optical communication network in the metropolis railroad accepts various kinds of control networks and data networks for train operation. As occasion demands, this paper presents effective application scheme and specialized organization about an optical communication operating system.

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RET Proto-Oncogene Mutation in Medullary Thyroid Carcinoma (갑상선 수질암 조직에서 RET 원암유전자의 돌연변이 양상)

  • Chung Woong-Youn;Song Hyeun-Jung;Cho Nam-Hoon;Park Cheong-Soo
    • Korean Journal of Head & Neck Oncology
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    • v.18 no.1
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    • pp.3-10
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    • 2002
  • Background: The molecular pathogenesis of hereditary medullary thyroid carcinoma is well known to be associated with germ-line mutation in the RET proto-oncogene and sporadic medullary thyroid carcinoma has been shown to carry somatic RET mutation especially in exon 13 and 16. The aim of this study is to evaluate the genetic background in the pathogenesis of the sporadic medullary thyroid carcinoma which shows extremely high incidence in Korea. Materials and Methods: Direct DNA sequencing for RET exon 13 and 16, as well as immunohistochemistrical assay for a monoclonal RET antibody were performed from 20 cases of archival tissues of medullary thyroid carcinoma. Results: Monoclonal RET antibody with C-terminal epitope showed comparatively stronger expression in tumor cells than in normal tissues and immunoreactive area in the tumor was $66.0{\pm}40.1%$. Direct sequencing of RET exon 13 revealed 4 cases of mis-sense mutations in Codon 778, Codon 767, and both in Codon 768 and 778. One case showed a silent mutation (ACG-ACT) in RET exon 16 (Codon 926). Conclusions: The strong RET immunoreactivity of medullary thyroid carcinoma may suggest that there could be a genetic alteration in oncoprotein level. RET proto-oncogene mutation may be involved in the evolutional process of medullary thyroid carcinoma in the aspect of molecular basis.

Antioxidant and anti-inflammatory effects of the ethyl acetate fraction of the Agastache rugosa extract (배초향 에틸아세테이트 분획의 산화방지 및 항염증 활성)

  • Kim, Bomin;Han, Yeong Eun;Lee, Hwa Jin
    • Korean Journal of Food Science and Technology
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    • v.49 no.3
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    • pp.331-337
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    • 2017
  • To evaluate the antioxidant activity of hexane, ethyl acetate, and butanol fractions obtained from Agastache rugosa extract, we measured the total polyphenol levels, DPPH radical scavenging activity, and reducing power. The ethyl acetate fraction of A. rugosa (AREA) displayed high phenolic levels, potent DPPH radical scavenging effect, and powerful reducing power. In addition, we examined the ability of AREA to inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-activated BV-2 microglia. AREA suppressed NO production and inducible nitric oxide synthase (iNOS) expression and downregulated interleukin-6 (IL-6) mRNA level in LPS-stimulated BV-2 microglia. Furthermore, we detected rosmarinic acid in AREA by HPLC, which suggested that rosmarinic acid could be one of the bioactive materials responsible for the antioxidant and anti-inflammatory activities of AREA. These results suggested that AREA may be a good source of functional foods with antioxidant and anti-inflammatory activities.

Variation of Psychophysiological Characteristics Related with Human Errors during a Simple Pointing Task (단순 지적과업 중 인간과오 관련 심리생리학적 특성의 변화)

  • Lim, Hyeon-Kyo
    • Journal of the Korean Society of Safety
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    • v.24 no.3
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    • pp.71-78
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    • 2009
  • During a learning process, a human being is assumed to experience knowledge-based behaviors, rule-based behaviors, and skill-based behaviors sequentially if Rasmussen was right. If any psycho-physiological symptom to those different levels can be obtained, it can be useful as a measure whether a human being is fully trained and has gotten a skill in his work. Therefore, this study aimed to draw relationships between human performance measures and psycho-physiological measures while committing a computer-simulated pointing task by utilizing the power spectrum technique of EEG data, especially with the ratio of relative beta-to-alpha band power. The result showed that, during correct responses, the ratio came to stabilize as all the performance data went stable. However, response time was not a simple linear function of task difficulty level only, but a joint function of task characteristics as well as behavior levels. Comparing relative band power ratios from errors and correct responses, activated states of one's brain could be explained, and characteristics of the task could understood. To tell that of pointing task, correlations around C3, C4, P3, P4 and 01, 02 area were significant and high in correct response cases whereas most correlation coefficients went down in error cases standing for imbalance of psycho-motor functions. Though task difficulty was the only one factor that could influence on relative band power ratio with statistical significance, it should be comprehended to mean a different way of expression indicating task characteristics since at least error-some situation could be explained with the help of relative band power ratio that absolute band power failed.